Lung cancer is the number one cancer killer in the USA. The development of sensitive and noninvasive approaches for lung cancer early detection will reduce the mortality, is thus an important clinical goal. Toward this goal, we have identified a set of genetic signatures that can be detected in sputum, providing a potential noninvasive biomarker panel for the early detection of lung cancer (Clin. Cancer Res, AACR Breaking News). However, the application of the genetic biomarkers in the clinical settings is limited by the cellular heterogeneity of sputum, which typically includes more than 97% macrophages and neutrophils and only 1% bronchial epithelial cells. The newly developed magnetic cell sorting (MACS) allows up to 109 cells separated in a single process, thus can efficiently enrich rare cells from biological fluids for molecular genetic analysis. The objective of this R03 study is to develop a noninvasive diagnostic test by detecting the genetic biomarkers in enriched bronchial epithelial cells from sputum for the early detection of lung cancer with high sensitivity and specificity. We propose to: 1) concentrate and purify bronchial epithelial cells from sputum by using MACS with antibody-beads to specifically deplete macrophages and neutrophils, and 2) determine diagnostic performance of the biomarker panel for the early detection of lung cancer in the enriched sputum samples from patients with stage I lung cancer. At completion, we will develop a genetic approach for effectively detecting the biomarker panel in sputum that can be used for the early detection of lung cancer with high sensitivity. This R03 research will set the stage for future large-scale cohort and multicenter studies designed to validate its utility for adoption in routine clinical settings. Resources that will contribute towards our objective include close collaboration of basic and clinical investigators with complementary expertise in molecular genetics, cytopathology, oncology, and biostatistics. Furthermore, an approved Institutional Review Board protocol will allow us to collect sputum samples from early stage lung cancer patients and controls, which will secure appropriate analysis and verify clinical applicability of the developing test in the project.